Join analysis MeDIP-seq / RNA-seq

Rene Welch

April 6, 2017

MeDIP-seq aligment rates

A more general view

A bit of processing

  • Removed the samples with less than 10M mapped reads

  • For each cell / treatment block pooled the 3 input replicates

  • For each gene in RNA-seq study, define the promoter regions as 500 bps upstream and 1K bps downstream of the TSS.

  • Count the number of extended reads (by 300 bps) in each promoter region and divide by sequencing depth

  • Fill matrix with \(\log_2 \left( \text{sample} / \text{input} \right)\)

MeDIP-seq heatmaps

Top: mC, Bottom: hmC

Left: benCaFBS, Middle: benMC, Right: ScottMC

Differential methylation analysis

  • From the heatmaps, we can observe there is not obvious differential methylation (for either treatment of cell lines)

  • We have a count matrix already: For each gene we counted the number of extended reads (by 300 bps) in the associated promoter region

  • We ran DESeq2 using the count matrices and evaluated the EBV vs NOKS tests

P.value diagnostics (MC)

P.value diagnostics (HMC)

Some notes

  1. With the HMC samples, possibly more genes are differentially methylated. I am going to focus on those gene lists.

  2. There are not a lot of differentially expressed genes, for example counting the differentially methylated genes (with HMC) for the Scott MC list, we get:

## # A tibble: 5 × 4
##   threshold ngenesDiffM ngenesDiffE ngenesInt
##       <dbl>       <dbl>       <dbl>     <dbl>
## 1     0.001           6         531         0
## 2     0.010          13        1206         0
## 3     0.050          73        2455        13
## 4     0.100         134        3369        26
## 5     0.300         357        5939       128
  1. Even less of these genes are differentially expressed too.

Methylated and expressed genes

Methylated and expressed genes

Genes in the green zone

Genes in the green zone

Genes in the green zone

Genes in the green zone

Genes in the green zone

Genes in the green zone

Genes in the red zone

Genes in the red zone

Genes in the red zone

Genes in the red zone

Genes in the red zone

Genes in the red zone

Genes in the red zone

Thanks